Environmental exposure to transition metals is linked to pathological processes of various neurodegenerative conditions since metal neurotoxicity often augments key degenerative changes including ionic imbalance, oxidative stress and protein aggregation. Several metal binding proteins regulate intracellular metal homeostasis and thereby maintain normal cellular function. Emerging evidence indicates that prion proteins are metal binding proteins that can efficiently bind to certain divalent cations including copper and manganese at the octapeptide repeat regions of the protein. Therefore, dysregulation of metal homeostasis has been suggested to play a role in the pathogenesis of prion diseases. Recent observations of elevated manganese (Mn) levels in the brain and blood of humans and animals afflicted with prion diseases suggest that manganese neurotoxicity may play a role in the etiology of prion diseases. Recently, we demonstrated that normal prion protein effectively attenuates manganese transport into neuronal cells and protects against manganese-induced oxidative stress, mitochondrial dysfunction, cellular antioxidant depletion, and apoptosis. While investigating these mechanisms, we unexpectedly found that manganese treatment upregulates cellular prion levels independent of transcription. Furthermore, we found manganese increases stability, suggesting that prion protein may promote the conversion of normal prion protein (PrPC) to the pathological form of prion (PrPSc), which results in the loss of normal prion protein's protective function against manganese neurotoxicity. Thus, the central hypothesis of this proposal is that manganese binds to the octapeptide (PHGGGWGQ) domain of cellular prion protein to increase the stability and accumulation of the protein. Manganese-induced stabilization of prion protein accelerates conformational conversion of PrPC to proteinase-resistant prion protein (PrPSc) aggregates and thereby induces neurotoxicity. This novel hypothesis will be tested through a systematic investigation of the following specific aims: i) to determine whether chronic exposure to manganese increases prion protein accumulation in animal models, ii) to determine the role of octapeptide repeat sequences in the manganese-induced stabilization of prion protein, iii) to determine whether chronic manganese exposure accelerates the accumulation and aggregation of the scrapie form of prion protein (PrPSc) and causes increased neuronal damage in a mouse model of prion disease, iv) to compare the effect of manganese on the accumulation and aggregation of PrPSc and on neuronal damage in mouse scrapie-infected prion overexpressing and octapeptide deletion transgenic animals (Tg20 and TgPrPDOR transgenic mice). Together, results from the proposed studies will not only provide new insights into the role of prion protein in manganese neurotoxicity but also will advance understanding of the role of metals in the pathogenesis of prion diseases.